The ultimate goal of these studies is to determine the role of intestinal peptide hydrolases in human nutrition and what role, if any, deficiencies of these enzymes play in human diseases. Substrates highly discriminating for the 5 peptide-hydrolyzing enzymes recently identified in human intestinal brush border will be used to determine the levels of these enzyme activities in intestinal biopsies of normal subjects and of patients with various intestinal diseases of undetermined etiology. Protein-separating techniques including ion-exchange chromatography, acrylamide gel electrophoresis, and isoelectric focusing will be used to separate and identify each peptide hydrolase present in the cytosol of human intestinal mucosa. Biochemical characteristics and substrates specificity of each isolated enzyme will be determined. Particular emphasis will be placed on determining substrates which are highly discriminating for each enzyme isolated. Various protein-separating techniques including immunoaffinity chromatography will be employed to purify the human intestinal brush border peptide hydrolyzing enzymes. Careful substrate specificity studies of each purified enzyme will be made.